Method of treatment of eyelid laxity

ABSTRACT

A method of treating eyelid laxity in a subject comprising exposing a tarsal plate of an eye, and applying to at least part of the exposed tarsal plate a photosensitizer that initiates crosslinking in response to photo-activating radiation; and irradiating the exposed tarsal plate with photo-activating radiation to initiate crosslinking in the tarsal plate tissue.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims priority to Australian Provisional ApplicationNo. 2017900561 entitled “METHOD OF TREATMENT” filed on 21 Feb. 2017, theentire content of which is incorporated herein by reference.

FIELD

The methods, apparatuses and compositions herein relate generally to amethod of treatment of eyelid laxity, and in particular to a methodcomprising exposure of the tarsal plate followed by irradiation in thepresence of a photosensitizer to enhance the rigidity or strength of thetarsal plate and inhibit the progression of the disorder.

BACKGROUND

The reference in this specification to any prior publication (orinformation derived from it), or to any matter which is known, is not,and should not be taken as an acknowledgment or admission or any form ofsuggestion that that prior publication (or information derived from it)or known matter forms part of the common general knowledge in the fieldof endeavor to which this specification relates.

The condition known as eyelid laxity (or lax eyelid), also known asfloppy eyelid syndrome (FES), is regarded as a subset of the conditionsgenerated by the laxity of the eyelid [see A. M. Fowler, J. J. Dutton:“Floppy eyelid syndrome as a subset of lax eyelid conditions:Relationships and clinical relevance (an ASOPRS thesis)”, Ophthal.Plast. Reconstr. Surg., 26 (2010) 195-204]. Eyelid laxity can be definedas an acquired hyperelasticity disorder where the inherent rigidity ofthe lids is lost. It can affect both upper and lower eyelids. Due tocorneal complications and associations with other eyelid disorders, afloppy eyelid can lead to blindness. Eyelid laxity is commonlyassociated with other conditions including obesity, cardiovasculardisease, obstructive sleep apnea, and eye rubbing.

The main clinical feature of a floppy eyelid is the extreme laxity ofthe upper or lower tarsus (or tarsal plate), which becomes pliant andrubbery. In normal conditions, the upper or lower tarsal plate is aspecialized dense fibrous tissue that is able to display significantrigidity and provide structural integrity, shape and firmness to theentire eyelid. Tarsal plate tissue can be regarded as a unique tissuewith transient character possessing features common to both cartilageand fibers [see a description of fibrocartilage tissues in M. Benjamin,J. R. Ralphs: “Biology of fibrocartilage cells”, Int. Rev. Cytol., 233(2004) 1-45]. The tarsal tissue consists mainly of fibrillar collagenstype I, III and, to a lesser extent, type VI, as well as elasticnetworks of fibrillin and elastin fibers [see: S. Milz, J. Neufang, I.Higashiyama, R. Putz, M. Benjamin: “An immunohistochemical study of theextracellular matrix of the tarsal plate in the upper lid in humanbeings”, J. Anat., 206 (2005) 37-45; D. G. Ezra, M. Beaconsfield, R.Collin: “Surgical anatomy of the upper eyelid: old controversies, newconcepts”, Expert Rev. Ophthalmol., 4 (2009) 47-57]. In the tarsalplates of the lax eyelids, the collagen expression, structure anddistribution remains the same as in normal tarsus, as was reported forinstance in: R. Goldberg, S. Seiff, J. McFarland, K. Simons, N. Shorr:“Floppy eyelid syndrome and blefarochalasis”, Am. J. Ophthalmol., 102(1986) 376-381; and E. Arrocker-Mettinger, R. Haddad, K. Konrad, F. J.Steinkogler: “Floppy eyelid syndrome: Investigations by light andelectron microscopies” (Ger.), Klin. Mbl. Augenheilk., 188 (1986)596-598. However, the decrease in the amount of elastin and matureelastic fibers observed histopathologically in the tarsal plate of a laxeyelid [as reported for instance in: P. A. Netland, S. P. Sugrue, D. M.Albert, J. W. Shore: “Histopathologic features of the floppy eyelidsyndrome. Involvement of tarsal elastin”, Ophthalmology, 101 (1994)174-181; U. Schlötzer-Schrehardt, M. Stojkovic, C. Hofmann-Rummelt, C.Cursiefen, F. E. Kruse, L. M. Holbach: “The pathogenesis of floppyeyelid syndrome. Involvement of matrix metalloproteinases in elasticfiber degradation”, Ophthalmology, 112 (2005) 694-704] is not consistentwith an enhanced elasticity and reduced mechanical strength that havebeen evidenced clinically. A more recent study [D. G. Ezra, J. S. Ellis,C. Gaughan, M. Beaconsfield, R. Collin, C. Bunce, M. Bailly, P. Luthert:“Changes in tarsal plate fibrillar collagens and elastic fiber phenotypein floppy eyelid syndrome”, Clin. Exp. Ophthalmol., 39 (2011) 564-571]has demonstrated that instead of a reduction of elastic fibers in thetarsal plates belonging to lax eyelids, the main process consists ratherof a change in the phenotype of these fibers. The same study alsoconfirmed that the expression and periodicity of the main fibrillarcollagens remain normal.

Current treatment methods for eyelid laxity include nocturnal lidshielding, taping or lubrication. These treatment methods generally onlyprovide temporary benefit. Surgical methods may be used to tighten thelid. These surgical methods include full-thickness wedge excision;lateral tarsal strip; lateral canthal tendon plication; or lateraltarsorrhaphy (D. G. Ezra, M. Beaconsfield, R. Collin: “Floppy eyelidsyndrome: Stretching the limits”, Surv. Ophthalmol., 55 (2010) 35-46).

These surgical horizontal tightening procedures usually only providetemporary benefit as the eyelid tissue continues to stretch with timeand the eyelid laxity returns. Furthermore, these surgical proceduresare usually performed under general anesthesia. This is a particularproblem as many patients with floppy eye syndrome are commonly alsoafflicted by cardiovascular disease, obesity and sleep apnea: conditionsthat require particular considerations with respect to administration ofgeneral anesthesia.

There is a need for improved methods for treating or inhibiting theprogression of eyelid laxity that overcome one or more drawbacks of thepresent therapies.

SUMMARY

The methods, compositions and apparatuses herein are predicated in parton the surprising discovery that exposure of a tarsal plate to radiationin the presence of a photosensitizer initiates crosslinking within thetissue, and particularly crosslinking of collagen in the tarsal platetissue. This crosslinking has been found to improve or restore strengthor rigidity to the tarsal plate resulting in arresting or slowing theprogression of eyelid laxity.

Accordingly, in one aspect the methods herein advantageously provide amethod of treating eyelid laxity in a subject comprising:

exposing a tarsal plate of an eye;

-   -   applying to at least part of the exposed tarsal plate a        photosensitizer that initiates crosslinking in response to        photo-activating radiation; and    -   irradiating the exposed tarsal plate with photo-activating        radiation to initiate crosslinking activity in the tarsal plate        tissue.

The method suitably further comprises making an incision to expose thetarsal plate and subsequent closure of the incision after irradiationwith the photo-activating radiation.

The inventors have discovered that it is desirable that thephotosensitizer is in solubilized form, suitably in the form of anaqueous composition. The method may advantageously include delivery ofO₂ at the site of crosslinking.

The methods herein may be applied to one or both eyes of a subject,individually, simultaneously or sequentially; or to the upper and/orlower tarsal plates of an eye, individually, simultaneously orsequentially. A method may be performed simultaneously or sequentiallyin combination with an additional surgical tightening procedure.

In another aspect, there is provided a system for crosslinking tarsalplate tissue comprising:

an applicator that applies or delivers a photosensitizer to an exposedtarsal plate of an eye; and

-   -   a radiation source that provides photo-activating radiation to        the tarsal plate with a beam profile of greater than 12 mm at a        distance of about 10 mm from a surface of the tarsal plate.

In another aspect, there is provided a kit or commercial package forcrosslinking tarsal plate tissue comprising:

an applicator that applies or delivers a photosensitizer to an exposedtarsal plate of an eye; anda radiation source for providing photo-activating radiation to thetarsal plate;together with instructions to treat eyelid laxity or crosslink a tarsalplate or tarsal plate tissue.

The system, kit or commercial package may further include a means ofdelivering O₂ to the site of crosslinking.

In a yet further aspect, there is advantageously provided herein aphotosensitizer that initiates crosslinking in response tophoto-activating radiation for use in treatment or prevention of eyelidlaxity.

In a yet further aspect, the methods herein advantageously provide useof a photosensitizer that initiates crosslinking in response tophoto-activating radiation for crosslinking of tarsal plate tissue.

In another aspect, there is provided the use of a photosensitizer thatinitiates crosslinking in response to photo-activating radiation for themanufacture of a medicament for treatment of eyelid laxity.

In another aspect, there is provided a use of a photosensitizer thatinitiates crosslinking in response to photo-activating radiation forinhibiting the progression or development of eyelid laxity in a subject.

The inventors have found it desirable to use riboflavin or apharmaceutically acceptable salt, derivative and/or solvate thereof as aphotosensitizer in combination with UV-A photo-activating radiation. Inpreferred embodiments the riboflavin is in a water soluble form, forexample a water soluble salt, derivative and/or solvate thereof.

In a further aspect, there is advantageously provided a pharmaceuticalcomposition comprising a photosensitizer, for example riboflavin or apharmaceutically acceptable salt, derivative or solvate thereof, for usein combination with photo-activating radiation for treatment orprevention of eyelid laxity.

In another aspect, the apparatuses herein also provide a kit orcommercial package comprising a pharmaceutical composition comprising aphotosensitizer that initiates crosslinking in response tophoto-activating radiation; together with instructions for use incombination with photo-activating radiation to treat eyelid laxity, orto effect crosslinking in a tarsal plate or tarsal plate tissue. In someembodiments the photosensitizer is riboflavin or a pharmaceuticallyacceptable salt, derivative or solvate thereof. In some embodiments thepharmaceutical composition is an aqueous composition. In someembodiments the photosensitizer is in solubilized form.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a representation of a horizontal skin incision on an uppereyelid.

FIG. 2 shows cauterization made with a Bovie hot-temperature cauterymachine through the underlying orbicularis oculi muscle down to theorbital septum.

FIG. 3 shows the creation of a pretarsal pocket in the central uppereyelid using the Bovie cautery machine.

FIG. 4 shows the insertion of a neurosurgical patty, soaked in aphotosensitizer solution, into the pretarsal pocket.

FIG. 5 shows the insertion of a corneal protector against the eye globe.

FIG. 6 shows the exposure of the anterior aspect of the tarsal plate ofthe upper eyelid, where irradiation is delivered.

FIG. 7 shows the closure of the incision using sutures.

FIG. 8 shows the stress-strain plots for untreated and UV-crosslinked(365 nm, 6 mW/cm²) sheep tarsal plates.

FIG. 9 shows comparative bar graphs of the experimentally measuredYoung's moduli, prior and after crosslinking with UV-A radiation at highirradiances of the sheep tarsal tissue from upper and lower eyelids. Theasterisk indicates statistical significance.

FIG. 10 shows comparative bar graphs of the experimentally measuredstress at 20% strain, prior and after crosslinking with UV-A radiationat high irradiances of the sheep tarsal tissue from upper and lowereyelids. An asterisk indicates statistical significance.

FIG. 11 shows comparative bar graphs of the experimentally measuredYoung's moduli, prior and after crosslinking of the sheep tarsal tissuefrom upper and lower eyelids with UV-A radiation at an irradiance of 45mW/cm², with the photosensitizer solution being applied either by directimmersion or through contact with fluid-saturated surgical patties. Anasterisk indicates statistical significance.

FIG. 12 shows comparative bar graphs of the experimentally measuredstress at 20% strain, prior and after crosslinking of the sheep tarsaltissue from upper and lower eyelids with UV-A radiation at an irradianceof 45 mW/cm², with the photosensitizer solution being applied either bydirect immersion or through contact with fluid-saturated surgicalpatties. An asterisk indicates statistical significance.

DETAILED DESCRIPTION 1. Definitions

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by those of ordinary skillin the art to which the methods, compositions and apparatuses hereinbelong. Although any methods and materials similar or equivalent tothose described herein can be used in the practice or testing of thepresent methods, compositions and apparatuses, preferred methods andmaterials are described. For the purposes of the methods, compositionsand apparatuses herein, the following terms are defined below.

The articles “a” and “an” are used herein to refer to one or to morethan one (i.e. to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element.

By “about” is meant a quantity, level, value, number, frequency,percentage, dimension, size, amount, weight or length that varies by asmuch 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1% to a referencequantity, level, value, number, frequency, percentage, dimension, size,amount, weight or length.

As used herein, the term “and/or” refers to and encompasses any and allpossible combinations of one or more of the associated listed items, aswell as the lack of combinations when interpreted in the alternative(or).

The phrase “aqueous carrier” is used herein to refer to a liquid aqueousdiluent, wherein the aqueous carrier includes, but is not limited to,water, saline, aqueous buffer and aqueous solutions comprising watersoluble or water miscible additives such as glucose or glycerol. Theaqueous carrier may also be in the form of an oil-in-water emulsion.

Throughout this specification and the claims which follow, unless thecontext requires otherwise, the word “comprise”, and variations such as“comprises” and “comprising”, will be understood to imply the inclusionof a stated integer or step or group of integers or steps but not theexclusion of any other integer or step or group of integers or steps.Thus, the use of the term “comprising” and the like indicates that thelisted integers are required or mandatory, but that other integers areoptional and may or may not be present. By “consisting of” is meantincluding, and limited to, whatever follows the phrase “consisting of”.Thus, the phrase “consisting of” indicates that the listed elements arerequired or mandatory, and that no other elements may be present. By“consisting essentially of” is meant including any elements listed afterthe phrase, and limited to other elements that do not interfere with orcontribute to the activity or action specified in the disclosure for thelisted elements. Thus, the phrase “consisting essentially of” indicatesthat the listed elements are required or mandatory, but that otherelements are optional and may or may not be present depending uponwhether or not they affect the activity or action of the listedelements.

As used herein, the terms “condition” or “disease” refers to anabnormality in the physical state of the body as a whole or one of itsparts.

As used herein, the term “salts”, “derivative” and “solvate” include anypharmaceutically acceptable salt, derivative, or solvate or any othercompound which, upon administration to the recipient, is capable ofproviding (directly or indirectly) a desired photosensitizer. Suitablepharmaceutically acceptable derivatives include esters, such asphosphate esters. Suitable pharmaceutically acceptable salts includesalts of pharmaceutically acceptable inorganic acids such ashydrochloric, sulfuric, phosphoric, nitric, carbonic, boric, sulfamicand hydrobromic acids, or salts of pharmaceutically acceptable organicacids such as acetic, propionic, butyric, tartaric, maleic,hydroxymaleic, fumaric, citric, lactic, mucic, gluconic, benzoic,succinic, oxalic, phenylacetic, methanesulfonic, toluenesulfonic,benzenesulfonic, salicylic, sulfanilic, aspartic, glutamic, edetic,stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic andvaleric acids. Base salts include, but are not limited to, those formedwith pharmaceutically acceptable cations, such as sodium, potassium,lithium, calcium, magnesium, ammonium and alkylammonium, particularlysodium. Also, basic nitrogen-containing groups may be quaternized withsuch agents as lower alkyl halides, such as methyl, ethyl, propyl andbutyl chlorides, bromides and iodides; dialkyl sulfates such as dimethyland diethyl sulfate; and others. Pharmacologically acceptable solvatesare known in the art, and include hydrates and alcoholates. Suitably,pharmaceutically acceptable solvates include hydrates, for examplemonohydrates, dihydrates and trihydrates. The skilled person willunderstand that a photosensitizer may be in the form of apharmaceutically acceptable salt, and/or a solvate and/or a derivative,for example riboflavin 5′-phosphate monosodium salt dihydrate and thelike. The preparation of salts, derivatives and solvates can be carriedout by methods well known in the art.

As used herein, the phrase “solubilized form” refers to a form where acompound, such as a photosensitizer, is dissolved in a liquid such thata solution comprising a uniform distribution of the compound is obtainedwhich is substantially free of solid compound. In some embodiments, theliquid is an aqueous carrier as described herein.

As used herein “water soluble form” refers to a chemical and/or physicalform of a compound, such as a photosensitizer, where the compound or asalt and/or derivative and/or solvate and/or polymorph thereof hassufficient solubility in water at ambient temperature to achieve aconcentration of from 0.1% to 20% w/v, 0.1% to 10% w/v, 0.1% to 5% w/v,0.1% to 3%, 0.1% to 2.5%, 0.1% to 1%, 0.1% to 0.5%, or 0.1% to 0.25%.Solubility can be determined using methods well known in the art.

The term “subject” or “individual” as used herein refers to a vertebratesubject, particularly a mammalian subject, for whom therapy orprophylaxis is desired. Suitable subjects include, but are not limitedto, primates; livestock animals such as sheep, cows, horses, deer,donkeys and pigs; laboratory test animals such as rabbits, mice, rats,guinea pigs and hamsters; companion animals such as cats and dogs; andcaptive wild animals such as foxes, deer and dingoes. In particularembodiments, the subject is a human. However, it will be understood thatthe aforementioned terms do not imply that symptoms are present.

The term “eyelid laxity” or “floppy eyelid syndrome” when used hereinrefers to conditions where an acquired hyperelasticity disorder in aneyelid has resulted in the inherent rigidity of the eyelid being lost.

When used herein, the term “photosensitizer” refers to a molecule that,on irradiation by photo-activating irradiation, produces a chemicalchange in another molecule through a photochemical process. Examples of“another molecule” include, for example, a crosslinker or crosslinkingagent such as O₂. A photosensitizer may convert O₂ molecules from thenormal O₂ triplet state to a more energetic singlet state that caninitiate crosslinking, for example in tissue molecules ormacromolecules. Further examples of “another molecule” include tissuemolecules or macromolecules, including collagen macromolecules. Aphotosensitizer, after exposure to radiation and transition to a moreenergetic state, may also produce a chemical change in collagen and/orother tissue molecules and initiate or generate crosslinking in thetissue. The skilled person will appreciate that optimum results will beachieved when the selected photosensitizer absorbs radiation at awavelength of the photo-activating radiation. The absorptionwavelength(s) of a photosensitizer can be determined byUltraviolet/Visible (UV/VIS) Spectrophotometry using a commerciallyavailable UV/VIS spectrophotometer in accordance with well knownprocedures. A photosensitizer will preferably be pharmaceuticallyacceptable, non-irritant and non-toxic.

When used herein, the term “crosslinker” or “crosslinking agent” refersto a chemical moiety that can chemically join two or more molecules, forexample by covalent bonding or ionic bonding, preferably by covalentbonding. An example of a crosslinking agent is O₂ which acts as acrosslinking agent when in the form of its high energy singlet state. Acrosslinker or crosslinking agent will preferably be pharmaceuticallyacceptable. A molecule may be a macromolecule. Preferably a crosslinkeror crosslinking agent will be substantially non-irritant and non-toxic.

The term “photo-activating radiation” when used herein refers toradiation that can activate a photosensitizer to produce a chemicalchange in another molecule. Suitably the photosensitizer absorbsradiation at a wavelength of the photo-activating radiation. In someembodiments the radiation is UV-A radiation.

The terms “treat”, “treating” or “treatment” as used herein cover thetreatment of eyelid laxity, and includes: inhibiting the disease orcondition, i.e., arresting its development; relieving the disease orcondition, i.e., causing regression of the disease or condition; orrelieving the symptoms resulting from the disease or condition, i.e.,relieving pain or inflammation without addressing the underlying diseaseor condition.

Each embodiment described herein is to be applied mutatis mutandis toeach and every embodiment unless specifically stated otherwise.

2. Methods

The present methods are based, in part, on the identification thatirradiation of tarsal plate tissue in the presence of a photosensitizercan significantly enhance the mechanical rigidity and strength of thetarsal plate. Thus, the inventors conceived that treatment of eyelidlaxity or inhibition of the development of eyelid laxity may be achievedby irradiating an exposed tarsal plate with photo-activating radiationin the presence of a photosensitizer.

These surprising findings have enabled the development of a method oftreating eyelid laxity comprising subjecting an exposed tarsal plate tophoto-activating radiation in the presence of a photosensitizer tocrosslink tarsal tissue and thus strengthen the tarsal plate. This leadsto slowing or arresting of the progression of eyelid laxity. Thisprovides a simple and effective procedure for treatment of eyelidlaxity. The method addresses a major drawback of known surgicalinterventions where the benefit is temporary as the eyelid continues tostretch and the laxity returns. The method impacts on the underlyingpathology in that it prevents or arrests tarsal stretching. Furthermore,the method can provide sustained benefit from surgery when combined withadditional tightening procedures, such as surgical tightening proceduresincluding full-thickness wedge excision; lateral tarsal strip; lateralcanthal tendon plication; or lateral tarsorrhaphy. The method can alsobe used as a prophylactic treatment in at-risk patients before thesymptoms arise. Furthermore, the method does not significantly alter themacroscopic eyelid anatomy, so it can be performed prior to, incombination with, or subsequent to other eyelid operations. Anotheradvantage of the method is that it can be performed under localanesthesia, making it more acceptable to patient affected by the oftenassociated conditions of cardiovascular disease, obesity and sleepapnea.

Without being bound by theory or mode of operation, it is believed thatthe photosensitizer initiates crosslinking within the tarsal platetissue, particularly crosslinking of collagen macromolecules, whenexposed to photo-activating radiation. This results in increasedrigidity and strength of the tarsal plate. It is believed that thephotosensitizer absorbs radiation and is raised to an energeticallyexcited state, which further initiates reactions within the tarsal platetissue, particularly in constitutive collagen (Type I reactions) andwith O₂ (Type II reactions). Both types of reaction are thought to beable to generate covalent crosslinkages between the macromolecules inthe tarsal tissue, particularly crosslinkages between collagenmacromolecules.

It is believed that the collagen expression, structure and distributionremains the same in the tarsal plates of the lax eyelids as that foundin the normal tarsus. However, the decrease in the amount of elastin andmature elastic fibers observed histopathologically in the tarsal plateof a lax eyelid is not consistent with an enhanced elasticity andreduced mechanical strength that is evidenced clinically. Instead of areduction of elastic fibers in the tarsal plates of lax eyelids, themain process appears to involve a change in the phenotype of thesefibers. The expression and periodicity of the main fibrillar collagensremain normal. Since an unaltered integration of the elastic fibernetworks (imparting compliance and elasticity) with the collagen fibers(imparting tensile strength and rigidity) is critical for ensuring andmaintaining the normal mechanical properties of the tarsal plate, it isbelieved that the stiffening of the collagen fibrils in the tarsal plateof a lax eyelid can compensate for the loss of strength and restore therigidity of the entire plate, leading to the arrest or slowing of eyelidlaxity.

Accordingly, in view of the surprising finding that irradiation oftarsal plate tissue in the presence of a photosensitizer cansignificantly enhance the mechanical rigidity and strength of the tarsalplate, the methods herein advantageously provides a method of treatingeyelid laxity in an individual comprising the steps of:

-   -   exposing a tarsal plate of an eye;    -   applying to at least part of the exposed tarsal plate a        photosensitizer that generates crosslinking in the tarsal plate        tissue in response to photo-activating radiation; and    -   irradiating the exposed tarsal plate with photo-activating        radiation to initiate crosslinking in the tarsal plate tissue.

Suitably the methods may be applied to a tarsal plate of an uppereyelid, a lower eyelid, or to both eyelids, subsequently orsimultaneously. A method may be applied to eyelids of one or both eyesof a subject, individually, subsequently or simultaneously.

The skilled person, such as an oculoplastic surgeon, will be familiarwith methods and procedures for exposing a tarsal plate of an eye,particularly the anterior aspect of the tarsal plate. Suitably thetarsal plate may be exposed by making an incision line horizontallywithin the eyelid crease (for an upper eyelid), or approximately 3 to 5mm, for example 4 mm, from the margin in the lower eyelid. The incisionis then deepened through the underlying orbicularis oculi muscle down tothe orbital septum and creating a pretarsal pocket in the central eyelidarea by extending medially and laterally using, for example, a hottemperature cautery instrument, to expose the anterior aspect of thetarsal plate. Suitably the area of the tarsal plate exposed is an areabetween 25 mm by 10 mm and 15 mm by 5 mm, for example approximately 20mm by 8 mm, on the upper eyelid; or an area between 25 mm by 6 mm and 15mm by 2 mm, for example 20 mm by 3 mm, on the lower eyelid.

A photosensitizer is suitably pharmaceutically acceptable, substantiallynon-toxic and substantially non-irritant. The skilled person willreadily understand that different photosensitizers will absorbphotosensitizing radiation of specific wavelengths according to thechemical structure of the chromophore, and will be able to match thephotosensitizer to the appropriate wavelength of photosensitizingradiation. In some embodiments the photosensitizer absorbs radiation inthe ultraviolet region of the electromagnetic spectrum, preferably longwavelength ultraviolet radiation (UV-A radiation, wavelength from 320 to400 nm). In some other embodiments, the photosensitizer absorbsradiation at a wavelength of 495-570 nm (green light). Preferably thephotosensitizer is pharmaceutically acceptable, non-toxic andnon-irritant. Preferably the photosensitizer has regulatory approval forfood and/or drug use. In some embodiments the photosensitizer maycomprise one or more chemical entities. In some embodiments there may bea single photosensitizer molecule type present.

In some embodiments the photosensitizer comprises riboflavin or apharmaceutically acceptable salt, derivative or solvate thereof.Riboflavin is also known as vitamin B2, and has the IUPAC name7,8-dimethyl-10-[(2S,3S,4R)-2,3,4,5-tetrahydroxypentyl]benzo[g]pteridine-2,4-dione.Preferably riboflavin is in a water soluble form, for example as a watersoluble derivative, salt or solvate, such as an alkali metal salt, ofriboflavin 5′-phosphate. Preferably the riboflavin derivative, solvateor salt is non-toxic and non-irritant. In some embodiments thephotosensitizer comprises a sodium salt of riboflavin 5′-phosphate or apharmaceutically acceptable solvate thereof, such as riboflavin5′-phosphate monosodium salt. Preferably a solvate is a hydrate.

In some embodiments the photosensitizer is rose Bengal(4,5,6,7-tetrachloro-2′,4′,5′,7′-tetraiodofluorescein disodium salt).Preferably rose Bengal is used in conjunction with irradiation by greenlight, i.e. radiation wavelength 495-470 nm.

In some embodiments the photosensitizer is selected from lucigenin,acridine orange, Quantacure QTX, Lissamine green B, fluorescein,Brilliant blue G, triamcinolone, or trypan blue. In some embodiments thephotosensitizer is selected from lucigenin, acridine orange, andQuantacure QTX(2-hydroxy-3-(3,4-dimethyl-9-oxo-9H-thioxanthen-2-yloxy)-N,N,N-trimethyl-1-propanaminiumchloride) and the irradiation has a wavelength greater than 300 nm(UV-A/visible).

In some embodiments, the photosensitizer is applied to at least part ofthe exposed tarsal plate. Suitably the photosensitizer is applied to atleast 25%, at least 50%, at least 75%, at least 80%, or at least 90% ofthe surface of the exposed tarsal plate. Preferably the photosensitizeris applied to substantially the entire surface of the exposed tarsalplate.

In some embodiments the photosensitizer is in solubilized form.Preferably the sensitizer is water soluble. Preferably the sensitizer isin aqueous solution. In preferred embodiments, the photosensitizer isformulated in an aqueous carrier. For example, the aqueous carrier maybe selected from, but is not limited to, saline, water, aqueous buffer,an aqueous solution comprising water and a miscible solvent, andcombinations thereof. In some embodiments, the aqueous carrier issaline. In preferred embodiments the photosensitizer and aqueous carrierform a sterile solution.

In some embodiments, the photosensitizer is formulated as an aqueousformulation, for example an aqueous solution, an aqueous gel, or an oilin water emulsion, preferably as an aqueous solution. In someembodiments the photosensitizer is present in an amount of 0.1 to 20%w/v. Suitable water soluble photosensitizers are readily available fromcommercial sources, for example riboflavin 5′-phosphate sodium salt,riboflavin 5′-phosphate sodium salt hydrate and riboflavin 5′-phosphatesodium salt dihydrate are available from Sigma Aldrich Co. LLC.Photosensitizers formulated as a solution are commercially available,for example ParaCel™ (Avedro, Inc, Waltham, Mass., USA) is acommercially available aqueous solution comprising 0.25% riboflavin inthe form of riboflavin 5′-phosphate sodium salt, and VibeX Rapid™(Avedro, Inc) is a commercially available solution comprising 0.1%riboflavin 5′-phosphate sodium salt. In some embodiments thephotosensitizer is commercially available as a sterile formulation in adispenser, for example as a sterile aqueous sodium riboflavin5′-phosphate solution packaged in a syringe, ampoule, vial or dropper,for example a single use syringe.

The skilled person will understand that the amount of photosensitizerrequired will depend on the identity of the photosensitizer. In someembodiments the amount of photosensitizer applied to the exposed tarsalplate is from 1 to 5 mL of a solution comprising from 0.1 to 0.5% w/v ofphotosensitizer, suitably 0.1 to 0.25% w/v. In some embodiments theamount of riboflavin applied to an exposed tarsal plate is 1.8 mL of0.25% riboflavin (as riboflavin 5′-phosphate) in aqueous carrier; or 2mL of 0.22% riboflavin (as riboflavin 5′-phosphate) in aqueous carrier;or 3 mL of 0.146% riboflavin (as riboflavin 5′-phosphate) in aqueouscarrier or 2 mL of 0.1% riboflavin (as riboflavin 5′-phosphate) inaqueous carrier.

The photosensitizer is applied to the exposed tarsal plate using anysuitable means known to the skilled person. In some embodiments thephotosensitizer is applied using an applicator. In some embodiments thephotosensitiser is applied dissolved in an aqueous carrier, preferablyas an aqueous solution. In some embodiments, the photosensitizer isinstilled directly onto the surface of the exposed tarsal plate. In someembodiments the photosensitizer is instilled onto the tarsal plate orpretarsal pocket using, a suitable applicator such as a syringe, pipetteor dropper. In some embodiments, a pretarsal pocket is formed surgicallyand the photosensitiser solution is instilled into the pretarsal pocket.In some embodiments, excess photosensitizer is removed from the tarsalplate or pretarsal pocket prior to irradiation. In some embodimentsexcess photosensitizer solution is removed from the tarsal plate usingabsorbent material, such as surgical sponge. In some embodiments,photosensitizer solution is first absorbed onto an absorbent material,preferably a disposable textile pad, for example a surgical sponge orneurosurgical patty, prior to application to the exposed tarsal plate orinsertion into the tarsal pocket. In some embodiments thephotosensitizer is applied to substantially the entire exposed surfaceof the tarsal plate.

In some embodiments the photosensitizer, preferably in the form of anaqueous solution, is allowed to remain in contact with the exposedtarsal plate for a period of from 6 to 60 minutes, for example 6 to 30minutes, 30 to 60 minutes, 10 to 30 minutes, 10 to 40 minutes, 6 to 15minutes, 6 to 20 minutes or 10 to 20 minutes. The exact timing willdepend on the amount and concentration of the photosensitizer, and canbe easily determined by the skilled person based on the concentration ofthe photosensitizer solution. In some embodiments the photosensitizerinstilled in the pretarsal pocket is 0.1% riboflavin (as riboflavin5′-phosphate, sodium salt) in aqueous solution which is allowed tocontact the tarsal plate for approximately 30 minutes. In someembodiments the photosensitizer instilled in the pretarsal pocket is0.25% riboflavin (as riboflavin 5′-phosphate, sodium salt) in aqueoussolution which is allowed to contact the tarsal plate for approximately30 minutes.

After the photosensitizer has contacted the tarsal plate for therequired length of time, the absorbent material, or excess solution, isremoved and the exposed tarsal tissue is exposed to radiation ofappropriate wavelength for the photosensitizer used. In some embodimentsthe photosensitizer is a sodium salt of riboflavin 5′-phosphate and theradiation is UV-A radiation of wavelength of about 320 nm to about 400nm. In some embodiments the UV-A radiation wavelength is approximately365 nm.

Treatment of the tarsal plate with photo-activating radiation may besimultaneous and/or subsequent to treatment with a photosensitizer. Insome embodiments additional photosensitizer may be applied to the tarsalplate during irradiation.

The skilled person will understand that the irradiation time necessaryto induce sufficient crosslinking in the tarsal plate tissue will bedependent on several factors including the irradiance intensitydelivered by the radiation source (mW/cm²), and the beam width.Preferably, use of an irradiance of about 3 mW/cm² up to about 150mW/cm² is envisaged. It will also be appreciated that the radiantexposure should not be detrimental to the health of the tissue. Aradiant exposure, or fluence, of about 4 to about 27 J/cm² is consideredappropriate. In some embodiments, the radiant exposure of the tarsaltissue, or fluence, is from about 5 to about 8 J/cm². In someembodiments, the fluence is about 27 J/cm². The skilled person will beable to determine the duration of the exposure required based on thepower of the radiation.

Suitably, the tarsal tissue is exposed to UV-A radiation (320 nm to 400nm, for example 365 nm) at an irradiance of about 3 to about 6 mW/cm²,for example about 3 mW/cm² or about 6 mW/cm². In some embodiments,irradiation of the photosensitizer treated tarsal plate with UV-Aradiation is maintained for a duration between about 6 minutes and aboutone hour, for example 6 to 40 minutes, 6 to 30 minutes, 6 to 20 minutesor 6 to 10 minutes.

In some embodiments, the irradiation may be carried out at highirradiance, for example at 30-45 mW/cm² for 1 to 4 minutes, for example2 to 3 minutes. In some embodiments the radiant exposure, or fluence, isfrom about 5 to 8 J/cm², for example from about 5.4 to 7.2 J/cm².

The skilled person will appreciate that there will be an upper limit tothe amount of irradiation that is considered useful. It is consideredthat irradiation levels of up to and including 150 mW/cm², for example150 mW/cm², may be used safely. Irradiation at a level of 150 mW/cm²should be carried out for up to 4 minutes, or up to 3 minutes; forexample 1 to 4 minutes, 1 to 3 minutes, 2 to 3 minutes, 1 to 2 minutes,or approximately 1 minute. Irradiances exceeding 150 mW/cm² areconsidered to be less effective in improving the stiffness or strengthof tarsal tissue. This is thought to be due to the presence of competingphotodegradative processes which may prevail over the beneficialcrosslinking reactions.

In some embodiments, the delivery of the irradiation is continuous. Insome embodiments, the delivery of irradiation is pulsed. In someembodiments, the radiation is applied using a beam profile of greaterthan 12 mm at 10 mm from a surface of the tarsal plate. In someembodiments, a narrower radiation beam of, for example, about 11 mm, maybe used if it is repositioned at time intervals, for example to ensurethat the entire surface of the tarsal plate is irradiated.

In some embodiments, the tarsal plate is irradiated over its entireexposed surface. In some embodiments the tarsal plate is irradiatedsubstantially where the photosensitizer has been applied.

In some preferred embodiments, an eye shield or protector, such as ametallic eye protector, is placed under the eyelid and against theanterior aspect of the eye globe prior to irradiation to protect the eyeglobe from radiation.

In some embodiments of the methods herein, irradiation is carried out inthe presence of one or more crosslinking agents. A crosslinking agentwill preferably be pharmaceutically acceptable. The skilled person willunderstand that a crosslinking agent will be preferably substantiallynon-irritant and non-toxic. In some embodiments irradiation is carriedout in the presence of O₂ gas. In some embodiments a method is carriedout in the presence of additional O₂ gas to provide O₂ concentrationsgreater than those present in the normal atmospheric conditions or inthe tarsal tissue. In some embodiments, the additional O₂ is provided atthe site of crosslinking during irradiation. In some embodiments O₂ gasis delivered to the surface of the tarsal plate in the proximity of theirradiation site, for example using a delivery device such as anapplicator. In some embodiments, the O₂ gas is humidified prior to use.Methods of generating humidified O₂ gas are known to the skilled person,and include passing the gas flow through a humidifier prior to deliveryto the required site.

In some embodiments, the methods herein comprise making an incision toexpose the tarsal plate and subsequent closure of the incision afterirradiation with the photo-activating radiation. Techniques for makingincisions are well known in the art and include, for example, use of ascalpel or a laser. Techniques and materials for effecting closure ofincisions are well known in the art, and include sutures, for examplesilk, catgut or synthetic sutures; adhesives, for example 2-octylcyanoacrylate; adhesive tapes or strips; or staples.

In some embodiments, a method may be performed simultaneously orsequentially in combination with an additional tightening procedure,such as a surgical tightening procedure. Suitable surgical tighteningprocedures are well known in the art, and include for examplefull-thickness wedge excision; lateral tarsal strip; lateral canthaltendon plication; or lateral tarsorrhaphy.

In some embodiments there is provided a method of treating eyelid laxityin an individual comprising the steps of:

-   -   (a) making an incision line within an eyelid crease in an upper        eyelid, and/or about 4 mm from the margin in a lower eyelid;    -   (b) anesthetizing the eyelid;    -   (c) sterilizing the eyelid;    -   (d) performing a horizontal skin incision on the eyelid on the        marked incision line;    -   (e) deepening the incision through the underlying orbicularis        oculi muscle down to the orbital septum;    -   (f) creating a pretarsal pocket in the central eyelid, extending        medially and laterally;    -   (g) exposing the anterior aspect of the tarsal plate;    -   and either:    -   (h) instilling a photosensitizer solution into the pretarsal        pocket; and    -   (i) removing the excess of solution after from 6-60 minutes;    -   or:    -   (j) as an alternative to step (h), inserting an absorbent        material soaked in an aqueous solution of photosensitizer into        the pretarsal pocket; and    -   (k) maintaining the absorbent material in the pretarsal pocket        for from 6-60 minutes; and removing the absorbent material;    -   and then:    -   (l) inserting an eye protector under the eyelid and against the        anterior aspect of the eye globe;    -   (m) applying directly to the exposed tarsal tissue a UV-A        radiation beam, optionally in the presence of O₂ gas delivered        close to the irradiation site to effect crosslinking, for        example for 1 to 60 minutes or 6 to 60 minutes, or 1 to 6        minutes, or 1 to 4 minutes, depending on the irradiance level;    -   (n) removing the corneal protector; and    -   (o) closing the skin incision on the eyelid.

In some embodiments there is provided a method of treating eyelid laxityin an individual comprising the steps of:

-   -   (a) making an incision line, suitably with an indelible skin        marker, within an eyelid crease in an upper eyelid, and/or 4 mm        from the margin in a lower eyelid;    -   (b) anaesthetizing the eyelid, for example by injecting local        anesthetic into the eyelid via transcutaneous or        transconjunctival delivery;    -   (c) sterilizing the eyelid skin, for example with Betadine, and        placing a sterile drape to expose the individual's face;    -   (d) performing a horizontal skin incision on the eyelid on the        marked line, for example using a scalpel;    -   (e) deepening the incision through the underlying orbicularis        oculi muscle down to the orbital septum, for example using a        hand-held hot-temperature cautery machine;    -   (f) creating with the hot-temperature cautery machine a        pretarsal pocket in the central eyelid, extending medially and        laterally;    -   (g) exposing the anterior aspect of the tarsal plate, preferably        approximately 20×8 mm on the upper eyelid, or 20×3 mm on the        lower eyelid, avoiding breach of the eyelid margin at the gray        line, and dehiscence of the levator aponeurosis;    -   (h) placing a traction suture in the pretarsal skin, close to        the incision line; and either:    -   (i) instilling a photosensitizer, for example an aqueous        solution of riboflavin 5′-phosphate monosodium salt into the        pretarsal pocket; and    -   (j) removing the excess of solution after, for example 6-60        minutes, using for example an absorbent material, such as a        sponge;    -   or:    -   (k) as an alternative to step (i), inserting an absorbent        material, such as a neurosurgical patty, soaked in an aqueous        solution of photosensitizer, such as riboflavin 5′-phosphate        monosodium salt, into the pretarsal pocket; and    -   (l) maintaining the absorbent material in the pretarsal pocket,        for example for 6-60 minutes and removing the absorbent        material;    -   and then:    -   (m) inserting a metallic eye protector under the eyelid and        against the anterior aspect of the eye globe;    -   (n) exposing the tarsal plate, for example by clamping the        traction suture to the drape, for example using an artery        forcep;    -   (o) applying directly to the exposed tarsal tissue a radiation        beam, for example UV-A, optionally in the presence of O₂ gas        delivered close to the irradiation site, for example for 1 to 60        minutes, or 6 to 60 minutes or 1 to 6 minutes, or 1 to 4        minutes, depending on the irradiance level;    -   (p) removing the corneal protector and the traction suture;    -   (q) checking the eyelid position, both open and closed, to        ensure the elevator palpebri or lower eyelid retractors are        functioning normally;    -   (r) closing the incision on the eyelid, for example with a        suture.

An exemplary procedure in accordance with the methods herein is carriedout in accordance with the method as illustrated in FIGS. 1 to 8. FIG. 1depicts the upper eyelid 11 and the incision 12 on the upper eyelid.FIG. 2 depicts cauterization through the muscles made with a hand-heldhot-temperature cautery machine, for example a Bovie cautery instrument21 (Bovie Medical Corporation, Purchase, N.Y., 10577, USA). FIG. 3depicts the creation of a pretarsal pocket 31 using the Bovie cauterytool 21. FIG. 4 depicts the insertion into the pretarsal pocket 31 of aneurosurgical patty 41 soaked in a photosensitizer solution. FIG. 5depicts the insertion of a metallic eye protector 51 under the eyelidand against the anterior aspect of the eye globe. FIG. 6 depicts theclamping of a traction suture 62 to the drape 61 in order to expose theanterior aspect of the upper tarsal plate, where the irradiation isapplied. FIG. 7 depicts the final closure of the incision 12 with suture71.

In one embodiment the method for the treatment of an eye affected byeyelid laxity is performed as follows. Local anesthetic (e.g. Alcaine)is applied to the patient's eye. An incision line is marked withindelible skin marker, within the eyelid crease in the upper eyelid, or4 mm from the margin in the lower eyelid. Local anesthetic (2.5-3 mL) isinjected into the eyelid either transcutaneously or transconjuctivally.The skin is sterilized with a suitable preparation (e.g. half-strengthBetadine™), and sterile drape is placed around surgical field, exposingthe patient's face. A horizontal skin incision 12 is performed on theeyelid 11 on the marked line, using a blade #15 scalpel, as shown inFIG. 1. As indicated in FIG. 2, the incision is extended through theunderlying orbicularis oculi muscle down to the orbital septum using ahand-held hot-temperature cautery machine, e.g. a Bovie cautery tool 21.A pretarsal pocket 31 is created in the central eyelid, extendingmedially and laterally, as shown in FIG. 3. The anterior aspect of thetarsal plate is exposed, approximately 20×8 mm on the upper eyelid, or20×3 mm on the lower eyelid. Care is taken not to breach the eyelidmargin at the grey line, and to avoid dehiscence of the levatoraponeurosis. A traction suture (for example using 6-0 Vicryl) is placedin the pretarsal skin, close to the incision line. Aqueous solution ofthe photosensitizer riboflavin 5′-phosphate monosodium salt (1 to 5 mLof 0.1 to 0.5% by weight) is instilled into the tarsal plate where itremains for from 6 to 60 minutes. Excess solution is removed by using asponge device, e.g. a Weck-Cell™ cellulose eye spear sponge.Alternatively, as depicted in FIG. 4, a neurosurgical patty (25×12 mm)41 trimmed to an appropriate size, is soaked for about 5 minutes in theaqueous solution of the photosensitizer, and then inserted into thepretarsal pocket 31, where it remains for from 6 to 60 minutes prior toremoval. A metallic eye protector 51 is inserted under the eyelid 11 andagainst the anterior aspect of the eye globe, as shown in FIG. 5. Thetraction suture 62 is clamped to drape 61 with an artery forcep, asshown in FIG. 6, in order to expose the tarsal plate and allow directirradiation of the tarsal tissue by a UV-A radiation beam. The UV-Aradiation may be provided by a commercially available UV cornealcrosslinking machine. The duration of the irradiation will depend on thewavelength and power of the radiation beam, and can be readilydetermined by the skilled person. Typical irradiation times at anirradiance of 3 mW/cm² are 6 to 60 minutes, suitably from 10 to 50minutes, 15 to 45 minutes, 20 to 40 minutes, 30 to 40 minutes, 20 to 30minutes or approximately 30 minutes. Typical irradiation times at anirradiance of 6 mW/cm² are 6 to 60 minutes, suitably from 10 to 40minutes, 15 to 45 minutes, 20 to 40 minutes, 20 to 30 minutes, 15 to 20minutes or approximately 30 minutes. Humidified O₂ gas can be deliveredat the site of crosslinking if desired. After the radiation treatment,the corneal protector and traction suture are removed. The eyelidposition is checked, in both opened and closed situations, to ensurethat the elevator palpebri or lower eyelid retractors are functioningnormally. If necessary, reattachment is performed surgically. The skinincision 12 on the eyelid 11 is then closed with the suture 71 (e.g. 6-0Vicryl, Ethicon Inc.), as shown in FIG. 7.

3. Systems and Apparatus

There is further advantageously provided a system for crosslinkingtarsal plate tissue comprising:

an applicator that applies or delivers a photosensitizer to an exposedtarsal plate of an eye; and

a radiation source for providing photo-activating radiation to thetarsal plate.

In another aspect there is provided a kit or commercial package forcrosslinking tarsal plate tissue comprising:

an applicator that applies or delivers a photosensitizer to an exposedtarsal plate of an eye; anda radiation source for providing photo-activating radiation to thetarsal plate;together with instructions to treat eyelid laxity or crosslink a tarsalplate.

In yet another aspect there is provided a kit or commercial package forcrosslinking tarsal plate tissue comprising:

an applicator that applies or delivers a photosensitizer to an exposedtarsal plate of an eye; anda radiation source for providing photo-activating radiation to thetarsal plate wherein the beam width is greater than 12 mm at 10 mm froma surface of the tarsal plate; and, optionally, instructions to treateyelid laxity or crosslink a tarsal plate.

In preferred embodiments, the radiation source provides a beam profileof greater than 12 mm at a distance of 10 mm from a surface of thetarsal plate, preferably the beam profile is from 13 mm to 15 mm at adistance of 10 mm from a surface of the tarsal plate.

Radiation sources are known in the art, and suitable sources arecommercially available. In some embodiments the radiation is UV-A with awavelength of from 320 to 400 nm. Radiation sources include thosesuitable for or intended for corneal crosslinking procedures such as,for example, XLink™ (Optos, Dunfermline, Scotland); CBM Vega XLinkCrosslinking System (Carleton Optical, Chesham, UK); LightLink CXL™(LightMed, San Clemente, Calif., USA); UV-X™ 2000 Crosslinking System(IROC Innocross, Zurich, Switzerland) and KXL™ CrossLinking System(Avedro Waltham, Mass., USA). These radiation sources typically have abeam width of approximately 10 to 12 mm, or about 11 mm. The skilledperson would understand that repositioning of such a radiation beamwould be necessary to effect radiation of the entire surface of anexposed tarsal plate. Alternatively, the radiation source instrument maybe modified to produce a greater beamwidth to irradiate substantiallyall of the tarsal plate surface at the same time.

Applicators for delivering or applying a photosensitizer to an exposedtarsal plate of an eye include syringes, droppers, vials, ampoules,pipettes, and absorbent materials such as surgical sponges andneurosurgical patties. In some embodiments a preferred applicator is anabsorbent material, such as a neurosurgical patty. The skilled personwill appreciate that absorbent materials may be cut or folded to sizewhere necessary.

A system, kit or commercial package as defined herein may also comprisea photosensitizer or a photosensitizer composition as defined herein.

The system, kit or commercial package may also comprise a deliverydevice that delivers O₂ gas to the exposed tarsal plate. In someembodiments the O₂ gas may be delivered using a nozzle, syringe,diffuser, or the like. Suitable methods of delivering O₂ to the exposedtarsal tissue and/or the site of irradiation or site of crosslinking aredisclosed in, for example, U.S. Pat. No. 8,574,277.

4. Compositions

Photosensitizers suitable for use in the methods described herein aremolecules having a chemical structure which includes a chromophore thatabsorbs radiation at a wavelength of the photo-activating radiation. Thephotosensitizer, on irradiation by photo-activating irradiation,produces a chemical change in another molecule, for example, O₂ and/orone or more tissue molecules, for example collagen molecules. This caninitiate crosslinking thus producing chemical change in the tissue. Insome embodiments the methods defined herein generate crosslinking incollagen. Preferably the photosensitizers are non-irritant, andpharmaceutically acceptable. A photosensitizer may be in the form of apharmaceutically acceptable salt, derivative or solvate thereof.Preferably a photosensitizer is soluble to a sufficient extent in anaqueous carrier to provide a concentration of from 0.1% to 10% w/v,preferably 0.1% to 5% w/v, 0.1% to 2% w/v, 0.1% to 1% w/v, 0.1% to 0.5%w/v, or 0.1% to 0.3% w/v. Suitable photosensitizers are known in theart, and are commercially available from, for example Sigma Aldrich Co.LLC. A preferred photosensitizer is riboflavin as an aqueous solubleform, for example a sodium salt of riboflavin 5′-phosphate, or apharmaceutically acceptable solvate thereof. Suitable photosensitizersare known in the art, and are commercially available from, for exampleSigma Aldrich Co. LLC).

Photosensitizer compositions used in the methods and systems definedherein are preferably pharmaceutical compositions and may be formulatedand administered using methods known in the art. Techniques forformulation and administration may be found in, for example, Remington:The Science and Practice of Pharmacy, Loyd V. Allen, Jr (Ed), ThePharmaceutical Press, London, 22^(nd) Edition, September 2012.

A photosensitizer composition is preferably in a form suitable fortopical administration to the tarsal plate. In some embodiments, thephotosensitizer composition is formulated for example as an emulsion,cream, lotion, gel, jelly, paste, ointment, solution, salve, orsolution, especially as a gel, oil in water emulsion or solution, asdescribed in, for example, the United States Food and DrugAdministration Monograph No. C-DRG-00201 entitled CDER Data StandardsManual Definitions for Topical Dosage Forms. In some embodiments, thecomposition is a solution.

Preferably the photosensitizer is in a solubilized form. Suitably, aphotosensitizer pharmaceutical composition is in a sterile aqueoussolution. In some embodiments the sensitizer is formulated in a sterileophthalmic solution. Suitably a sterile aqueous photosensitizer solutionis packaged in a vial, ampoule, syringe, for example as a single usesyringe.

Aqueous compositions, such as solutions, of a photosensitizer, forexample a water soluble form of riboflavin, are commercially availableor may be prepared using known methods. For example, a water solublesalt, derivative or solvate of riboflavin may be dissolved in an aqueouspharmaceutically acceptable carrier selected from, but not limited to,saline, water, aqueous buffer, an aqueous solution comprising water anda miscible solvent, and combinations thereof. An aqueous composition mayadditionally include pharmaceutically acceptable excipients selectedfrom viscosity modifiers such as dextran, buffers, rheology modifiers,surfactants, and chelating agents.

Commercially available riboflavin solutions include VibeX Xtra™, [0.22%riboflavin, saline isotonic]; MedioCROSS TE [0.25% riboflavin5′-phosphate, 1.2% HPMC (hydroxypropylmethylcellulose), 0.01% BAC(benzalkonium chloride)]; MedioCROSS M [0.1% riboflavin 5′-phosphate,1.1% HPMC]; VibeX Rapid™, [0.1% riboflavin 5′-phosphate, saline, HPMC];ParaCel™ [0.25% riboflavin 5′-phosphate, HPMC, BAC], Photrexa [0.146%riboflavin 5′-phosphate ophthalmic solution] or Photrexa viscous [0.146%riboflavin 5′-phosphate in 20% dextran ophthalmic solution], allavailable from Avedro Inc, USA. Riboflavin is typically in the form ofriboflavin 5′-phosphate sodium salt.

Materials for use in surgical procedures are well known to the skilledperson and are readily available from commercial sources. Suitable localanaesthetics for anaesthetizing the eyelid are well known to the skilledperson and include, for example Alcaine® (Proparacaine hydrochloride),Naropin® (Ropivicane hydrochloride), Marcaine™ (Bupivacainehydrochloride) and Novesin® (Benoxinate, Oxybuprocaine hydrochloride).Suitable antiseptics include Betadine® (povidone/iodine) and Betasept®(chlorhexidine). Wound closure may be effected using techniques known inthe art and include sutures; adhesives; adhesive tapes; staples and thelike.

In order that the methods, compositions and apparatuses may be readilyunderstood and put into practical effect, particular preferredembodiments will now be described by way of the following non-limitingexamples.

EXAMPLES Materials and Methods

The experiments described were performed on tarsal plates excised fromsheep, based on the fact that the ovine and human tarsal plates arestructurally and functionally identical, as shown in: M. J. Boileau, M.A. Gilmour: “Diseases of the eye”, in Sheep and Goat Medicine, 2nd Edn,D. G. Pugh, A. N. Baird (Eds), Elsevier, Amsterdam, 2012, pp. 406-410;D. A. Samuelson: “Ophthalmic structures”, in Essentials of VeterinaryOphthalmology, 3rd Edn, K. N. Gelatt (Ed.), Wiley, Oxford, U K, 2014,pp. 12-20.

Riboflavin and its salts, solvates and derivatives are readily availablefrom commercial sources. For example USP riboflavin 5′-phosphatemonosodium salt is commercially available from Sigma-Aldrich Co LLC.Aqueous 0.25% riboflavin is commercially available as ParaCel™ fromAvedro Inc, USA. Aqueous 0.1% riboflavin 5′-phosphate (dextran free) isavailable from Avedro Inc, USA as VibeX Rapid™.

Commercially available corneal crosslinking systems are available, forexample, Opto XLink™ corneal crosslinking system (Opto Electrônica S/A,Sao Carlos,—SP 13563-330, Brazil; Opto USA Corp, North Miami,Fla.-33181, USA; opto.com.br); or KXL Accelerated Cross-Linking System(Avedro, Inc, Waltham, Mass. O₂₄₅₁, USA; avedro.com). Alternatively, theUV curing system OmniCure® 1500 (Excelitas Technologies Corp, Waltham,Mass., O₂₄₅₁; excelitas.com) can be used to provide a large range ofirradiances.

Example 1 Crosslinking of Sheep Tarsal Plates

Upper and lower eyelids were isolated from cadaveric sheep eyes (1-2years old), and the tarsal plates were excised. The resulting tarsalstrips, with a mean thickness of 1.5±0.2 mm for both upper and lowereyelids, were treated with a commercially available riboflavin solution(ParaCel™) for 30 min, and then irradiated in a commercially availablecorneal crosslinking system (Opto XLink™) with UV-A radiation (365 nm)at an irradiance of either 3 mW/cm² or 6 mW/cm², for various durations.Humidified O₂ gas was delivered at the site of crosslinking.

Example 2 Tensile Measurements on Sheep Tarsal Plates

Tensile measurements on either untreated and crosslinked tarsal stripswere performed in an Instron mechanical microtester equipped with a 50-Nload cell, at a set gauge distance of 6 mm and a speed of 1.5 mm/min.The stress-strain plots were recorded and Young's moduli were computedin the linear region. Four to six measurements for each specimen wereperformed and recorded. The results were statistically processed by theone-way analysis of variance (ANOVA) in conjunction with Tukey-Kramermultiple comparisons, using the GraphPad® Prism software (Version 6.0).

Example 3 Stress-Strain Plots of the Sheep Tarsal Plates

FIG. 8 shows the stress-strain plots of an untreated tarsal plate and aUV-crosslinked tarsal plate, demonstrating the effect of thecrosslinking process on the mechanical properties of the tarsalfibrocartilage tissue.

Example 4 Mechanical Properties of Sheep Tarsal Plates

Table 1 presents the UV-irradiation conditions, and the values ofYoung's modulus and stress at 20% strain for the sheep tarsal stripsisolated from the upper and lower eyelids. The crosslinking processincreased the stiffness of the tarsal tissue, as indicated by the highervalues of Young's modulus and stress.

TABLE 1 Young's modulus and stress values at 20% strain measured for thesheep tarsal plates excised from upper and lower lids UV Increase inConditions Time Young's Young's Tarsus Irradiance (min- modulus modulusStress location (mW/cm²) utes) n (MPa) (%) (kPa) Upper Untreated — 6 3.1± 1.5 — 319.6 ± 224.6 eyelid 3 30 4 3.9 ± 0.6 25.8 449.6 ± 104.3 6 30 44.8 ± 1.1 54.8 480.1 ± 121.4 Lower Untreated — 6 4.2 ± 1.8 — 459.6 ±212.9 eyelid 3 30 4 5.0 ± 1.4 19.0 685.6 ± 327.2 6 30 4 5.5 ± 0.8 31.0477.6 ± 163.2

Example 5 Crosslinking of the Sheep Tarsal Plates at High Irradiance

Upper and lower eyelid tarsal strips excised from cadaveric sheep eyeswere crosslinked at high irradiance in a KXL Accelerated Cross-LinkingSystem machine, which delivered a maximum irradiance of 45 mW/cm² at themaximum fluence of 7.2 J/cm². The tarsal specimens were immersed in 0.1%riboflavin solution (VibeX Rapid™) for 30 min, and then irradiatedcontinuously with radiation UV-A (365 nm) without delivery of O₂ gas atthe site of crosslinking. The irradiance levels of 150 mW/cm² and 250mW/cm², respectively, were generated using the UV curing systemOmniCure® 1500, equipped with a 365-nm filter.

Table 2 presents the conditions used for crosslinking at highirradiances.

TABLE 2 Conditions for crosslinking at high irradiances IrradianceFluence Time Number of samples (n) (mW/cm²) (J/cm²) (s) Upper eyelidLower eyelid 30 5.4 180 2 2 45 5.4 120 3 3 45 7.2 160 3 3 150 27.0 180 35 250 45.0 180 3 5

FIGS. 9 and 10 show, respectively, the effect of irradiation at highirradiances on Young's modulus (stiffness) and on tensile stress(mechanical strength) of the tarsal tissue specimens excised from upperand lower eyelids of sheep eyes. The process of crosslinking increasedsignificantly the rigidity of the tarsal fibrocartilage tissue. However,at irradiance levels exceeding 150 mW/cm², the increase in stiffness orstrength of the tarsal tissue appears to cease as an opposite effect,potentially due to photodegradation, becomes evident leading to thedecline of these properties.

Example 6 Effect of Different Procedures for the Application ofPhotosensitizer Solution

Two different approaches were used for treating with 0.1% riboflavinsolution (VibeX Rapid™) the ovine upper and lower eyelid tarsal stripsprior to their irradiation. In the first procedure, the strips weresoaked in the riboflavin solution for two different durations (15 min or30 min), and then irradiated. In the second procedure, the strips werekept in contact with surgical patties that were completely soaked inriboflavin solution. The contact between the tarsal strips and pattieswas maintained for 15 min or 30 min. Following the riboflavin treatment,the samples were irradiated using the OmniCure® 1500 machine, whichdelivered a maximum irradiance of 45 mW/cm² at the maximum fluence of7.2 J/cm². The results presented in FIGS. 11 and 12 indicate that themechanical properties were enhanced when the photosensitizer solutionwas applied to the tarsal tissue through an absorbent material such as asurgical patty.

The disclosure of every patent, patent application, and publicationcited herein is hereby incorporated herein by reference in its entirety.

The citation of any reference herein should not be construed as anadmission that such reference is available as “Prior Art” to the instantapplication.

Throughout the specification the aim has been to describe the preferredembodiments of the methods, compositions and apparatuses withoutlimiting to any one embodiment or specific collection of features. Thoseof skill in the art will therefore appreciate that, in light of theinstant disclosure, various modifications and changes can be made in theparticular embodiments exemplified without departing from the scope ofthe methods, compositions and apparatuses defined herein. All suchmodifications and changes are intended to be included within the scopeof the appended claims.

What is claimed is:
 1. A method of treating eyelid laxity in a subjectcomprising: exposing a tarsal plate of an eye; applying to at least partof the exposed tarsal plate a photosensitizer that initiatescrosslinking in response to photo-activating radiation; and irradiatingthe exposed tarsal plate with photo-activating radiation to initiatecrosslinking in the tarsal plate tissue.
 2. The method according toclaim 1 wherein the photosensitizer is in solubilized form.
 3. Themethod according to claim 1 wherein the photosensitizer is riboflavin ora pharmaceutically acceptable salt, derivative or solvate thereof. 4.The method according to claim 1 wherein the photosensitizer isriboflavin 5′-phosphate or a pharmaceutically acceptable salt or solvatethereof.
 5. The method according to claim 1 wherein the photo-activatingradiation is UV-A radiation.
 6. The method according to claim 1 whereinthe method comprises: exposing a tarsal plate of an eye by making anincision; applying to at least part of the exposed tarsal plateriboflavin or a pharmaceutically acceptable salt, derivative or solvatethereof in solubilized form; irradiating the exposed tarsal plate withUV-A radiation to initiate crosslinking in the tarsal plate tissue; andclosing the incision.
 7. The method according to claim 1 wherein themethod further includes delivery of O₂ gas at a site of irradiation. 8.The method according to claim 1 performed either simultaneously orsequentially in combination with an additional tightening procedure. 9.A system configured to crosslink tarsal plate tissue comprising: anapplicator configured to apply a photosensitizer solution to an exposedtarsal plate of an eye; and a radiation source configured to providephoto-activating radiation to the tarsal plate wherein the radiationsource provides a beam profile of greater than 12 mm at 10 mm from asurface of the tarsal plate.
 10. The system of claim 9 furthercomprising a means for delivering O₂ to the tarsal plate.
 11. A methodof crosslinking tarsal plate tissue comprising topical application tothe tarsal plate of a photosensitizer that initiates crosslinking inresponse to photo-activating radiation and, simultaneously orsubsequently, treating the tarsal plate with photo-activating radiation.12. The method according to claim 11, wherein the photosensitizer is apharmaceutical composition comprising riboflavin or a pharmaceuticallyacceptable derivative, salt or solvate thereof.
 13. The method accordingto claim 12, wherein the pharmaceutical composition comprises an aqueoussolution of riboflavin 5′-phosphate or a pharmaceutically acceptablesalt or solvate thereof.
 14. The method according to claim 11, whereinthe radiation is UV-A radiation.
 15. A kit for performing the method ofclaim 1 comprising a pharmaceutical composition comprising aphotosensitizer that initiates crosslinking in response tophoto-activating radiation together with an applicator adapted to applythe photosensitizer to a tarsal plate.
 16. A kit according to claim 15wherein the photosensitizer is riboflavin or a pharmaceuticallyacceptable salt, derivative or solvate thereof.
 17. A kit according toclaim 16 wherein the riboflavin pharmaceutical composition is an aqueoussolution.